DESCRIPTION: This application proposes to study DNA binding by a number of single-stranded DNA binding proteins. This group includes two well- studied, prokaryotic proteins, the T4 gene 32 protein and the E. coli SSB, and two eukaryotic proteins, RP-A of yeast and D-SSB of Drosophila. Although the proteins T4 gp32 and E. coli SSB seem genetically unrelated, they resemble each other in their interaction with DNA. Dr. Prigodich's postdoctoral research included physical approaches to mapping functional domains in SSBs, most particularly T4 gene 32 protein. In spite of their lack of sequence-specificity, using single- stranded oligonucleotides, Dr. Prigodich has developed a method to footprint DNA with SSB. The method uses a 75-mer oligonucleotide dT and potassium permanganate. The permanganate specifically attacks the thymine bases unprotected by protein. The sugar-phosphate backbone of the oligonucleotides can then be cleaved at the site of thymine modification by piperidine treatments. The resulting fragments can be resolved by capillary gel electrophoresis. The fact that gene 32 protein interacts with its own RNA is recognized as an important and interesting aspect of its activity. It is proposed to use capillary electrophoresis with fluorescently tagged oligonucleotides in order to achieve higher resolution and greater sensitivity.